Mass Spectrometric Analysis of Thiocitrulline Bound Proteins Extracted from Human Keratinocyte Cells – Chemical Proteomics and Mass Spectrometry Experiments to Identify Proteins that Binds to Thiocitrulline


  1. Gebremedhin, M.G.
  2. Mah, D.C.W.
  3. Nelson, P.
  4. Chan, N.W.C.
Corporate Authors
Defence R&D Canada - Suffield, Ralston ALTA (CAN)
Previous studies have shown that some arginine analogue compounds, such as L-thiocitrulline, protect human keratinocytes against the toxicity of sulfur mustard (H). Their mechanism of action, however, is not yet known, but in the case of L-thiocitrulline there is reason to suspect that it binds to a cell membrane structure due to its speed of action. In this work we describe the development of proteomics-based approach to identify proteins that bind thiocitrulline using ESIMS. Membrane proteins were isolated from the first passage of human skin keratinocytes cells which were prepared from neonatal foreskins. Thiocitrulline-bound proteins were subjected to trypsin digestion. Tryptic-digested peptides were eluted and then desalted using SCX zip tip. Peptide separation was done on a C18 capillary column with direct infusion via nanospray ionization to a 3200 QTrap instrument. Independent data acquisition (IDA) files were used with both MASCOTTM 2.2 and ProteinPilot™ 2.0.1 search engines against the Swiss-Prot protein database. A total of 9 proteins were identified using both search engines. Six proteins were suspected of binding to thiocitrulline.

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Report Number
DRDC-SUFFIELD-TM-2008-245 — Technical Memorandum
Date of publication
01 Dec 2008
Number of Pages
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